中国生物工程杂志

Avian influenza, caused by avian influenza virus (AIV), is a highly contiguous disease. AIV can infect almost all wild and domestic poultries and cause severe economic losses for the poultry industry. The M2 protein of AIV is a transmembrane protein. It has highly conserved antigenic epitopes and is a potential candidate antigen for avian influenza vaccine with cross-protection. Marek's disease virus(MDV), avian herpesvirus have mostly been used as virus vectors to express a foreign gene for the protection of chickens against poultry. Among MD vaccine viruses, attenuated MDV1 strains such as CVI988 are more suitable for be vaccine vectors because they have the closest antigenic similarity to the very virulent(vv) MDV1. To develop a recombinant herpesvirus -based AI vaccine, the M2 gene of Avian Influenza virus (AIV) ligated with green fluorescent protein (EGFP) was inserted into the US gene, which located in the nonessential region of MDV1 (CVI988) genome and an EGFP marked recombinant plasmid was obtained. The M2 gene of Avian Influenza virus was expressed under the control of a CMV immediately early promoter. Through homologous recombination, this construct was transfected into cells infected with Marek's disease virus CVI 988/Rispens and the recombinant MDV1expressing AIV M2 gene was screened out. The PCR, dot-blot and western-blot results demonstrated that AIV M2 gene was not only cloned into the genome of MDV1 (CVI988) but also expressed. A group of one day-old chickens was injected with the recombinant virus and the specific antibody against M2 protein could be detected by ELISA after 21 days. When chickens were inoculated with commercial H9N2 subtype AI vaccine, the titers of antibody against AIV hemagglutinin from chickens inoculated with the recombinant MDV1were higher significantly (P<0.05) than those of H9N2 subtype AI vaccine alone. In addition, the excretive amount of live AIV from throat and cloacae of chickens inoculated with recombinant MDV1 were less significantly (P<0.01) after challenged with Avian influenza virus A/Chicken/Guangdong/00（H9N2）. All these results indicated that the recombinant MDV1expressing AIV M2 gene could be responsible for the prophylactic of avian influenza.